This invention relates to a procedure for the immobilization of micro-organisms and animal cells, in particular for anaerobic processes on porous, inorganic carrier bodies, to the subsequently obtained carrier bodies bearing a growth of micro-organisms, and to the carrier bodies suitable for immobilization, inter alia.
The immobilization of micro-organisms and cell material on solid bodies provides a means of providing such materials in abundance at a desired site. This is of significance in particular in the case of biotechnological processes. Both aerobic and anaerobic biotechnological processes should produce as high a space-time yield (substrate "turnover" per volume and time unit) as possible. This requirement can be met all the more readily, the greater the concentration of the active cells serving a production and catalyst role simultaneously.
High concentrations of cells are readily achieved in aerobic systems, where cell growth occurs virtually unhindered. In anaerobic systems, in contrast, cell growth is subjected to a limitation from the beginning, so that only relatively low biomass concentrations are achieved. In recent times, however, it is just these anaerobic systems that have attracted particular attention due to the favorable energy balance involved (e.g., biogas formation on the one hand, and no need for energy expenditure for the oxygen supply necessary for aerobic systems on the other). It has bene recognized that, using such systems and with little energy, it is often possible to produce valuable disproportionated products from inexpensive substrates. A particular example of this is the anaerobic treatment of highly concentrated waste water, resulting in the conversion of up to 95% of the organic contamination into biogas, with the simultaneous production of only 3-4% biomass. The low growth of micro-organisms in anaerobic systems makes it especially necessary to "hold back" and concentrate the biomass. This may also be of interest in the case of aerobic systems, for example, in the solution of separating problems.
For this reason, the immobilization of micro-organisms on solid carriers has long been practiced and investigated. In this connection, inexpensive, readily available carrier materials from the environment, such as sand, lava rocks, ceramics, activated charcoal, anthracite, glass, etc., have been investigated. With these, a more or less good immobilization of the micro-organisms can be accomplished More recently, more organic carrier materials have attracted interest: Thus, I. Karube et al (Biotechnol. Bioeng. Vol. 22 (1980), pages 847-857), describes a study of the immobilization of methane-producing bacteria on polyacrylamide gel, agar gel and collagen membranes, of which only the agar gel was found to be suitable. At the same time, however, attention was drawn to the low diffusion capacity of the nutrients and of methane through the agar gel. P. Scheter et al (Biotechnol. Bioeng Vol 30 23 (1981), pages 1057-1067) reported the immobilization of Methanosarcina barkeri on a Ca.sup.2+ - cross-linked alginate network, which was studied in the form of pellets with diameters varying between 1.2 and 3.7 mm. In this report, in contrast to that of P.S.J. Cheetham et al (Biotechnol. Bioeng. Vol. 21 (1979) 2155 ff.) who maintained that substrate transport into the alginate pellets is delayed, no difference in the activity of the micro-organisms dependent on the diameter of the pellets was found.
On the occasion of the 5th Symp. Techn. Mikrobiol. held in Sept. 82 in Berlin, B. Kressdorf et al reported on the immobilization of yeasts and bacteria by Ca-alginate gel. Comparative investigations with a variety of different types of carriers were described; cross-linked alginate microspheres of high solidity and a diameter of less than 1 mm bearing biomass, were said to be particularly useful.
Comparative investigations were also carried out by P. Huysman et al (Biotechn. Letters, Vol. 5 Nr. 9, (1983), pages 643-648). The carrier materials they studied were particles--about 5 mm in size--of sepiolite, zeolite, Argex (fire-expanded clay with surface pores of 0.1-7.5 .mu.m), and glass beads, all examples of "non-porous materials", and, as examples of "porous materials", natural sponge with a porosity of about 50% and pore sizes varying from the .mu.m range to the cm range, and non-cross-linked polyurethane foam with a porosity of about 30% and pore sizes varying from the .mu.m to the mm range, and, finally, various sorts of cross-linked polyurethane foams with a porosity of 97% and uniform pore diameters of (a) 2.21 mm; (b) 430 .mu.m and (c) 270 .mu.m. Finally, polyurethane foam,
coated with bentonite, with a uniform pore size of 430 .mu.m was also incorporated into the study. It was established that of the "non-porous materials", only sepiolite revealing on crystalographic examination fine bundles of needle-like crystals having a length of 2 .mu.m, permitted a useful formation of colonies. The bundles of "needles" revealed numerous gaps of a size corresponding to that of bacteria. The porous materials however, proved to be particularly suitable, the leading factor being found to be the great porosity and the size of the pores. In particular, the material having 430 .mu.m pores and a porosity of 97%, with and without a coating of bentonite, produced favorable results. With this cross-linked polyurethane foam material, within a period of 2 weeks, approximately 25 liters of biogas (65% methane) per liter of reactor a day were produced.
In DE-OS 28 39 580, a number of porous carrier materials, in particular glass frits, are indicated for the immobilization of micro-organisms, 70% or more of the pores of which are at least as large as the smallest dimension of the micro-organisms, but smaller than 4 to 5 times the greatest dimension (in yeast cells or bacteria). It was established that both non-porous boron silicate glass, and also glass frits with pores larger than 20 .mu.m in diameter, were appreciably poorer than material bearing pores of less than 20 .mu.m.
Despite the numerous different investigations into carrier materials, and the development of, in part very useful, carrier bodies, however, the problem of immobilization of micro-organisms has not yet been resolved completely satisfactorily. In individual situations, different aspects, such as density, abrasion resistance, stability, long-term behavior, wettability and the like, are problematical. Consequently, the general objective of a particularly high level of effectiveness of the immobilized biomass has not yet been achieved.